User Manual. pe-300 white. White Light Fluorescence Illumination System. DOC-005 Iss 4 1

Transcription

1 User Manual pe-300 white White Light Fluorescence Illumination System DOC-005 Iss 4 1

2 Table of Contents 1. Introduction 2. Safety 3. Getting started system components 4. Installation / set-up 5. Configuration of LEDs as a white Illumination System 6. Operation manual control 7. Remote Operation TTL and USB 8. Optical setup 9. Settings /additional information 10. Troubleshooting 11. Routine care and maintenance 12. Fitting pe-300 white to a different microscope 13. pe-300 white with liquid light guide option 14. Product specifications 15. Product options and order codes 16. Warranty & repairs 17. Contact details DOC-005 Iss 4 2

3 1 Introduction CoolLED s pe-300 white illumination system is designed to offer broad spectrum LED illumination for general use in fluorescence microscopy applications. It can be fitted directly to the microscope as a better and safer alternative to high pressure mercury or metal halide illuminators. Spectral coverage is from the UV (DAPI excitation) to the Red region (Cy5 excitation). It will excite common fluorophores used in hospital and research environments. With a comprehensive range of microscope adaptors, the pe-300 white can be fitted to most current and older microscopes. The result is a safe, convenient illumination system which will last for many years without any additional operating costs. This manual should give you all the information required to install and operate your new illumination system. Additional information can be found on our website at DOC-005 Iss 4 3

4 2 Safety Precautions While LEDs are a much safer illumination system than the mercury and metal halide lamps that they replace in the application of fluorescence microscopy, precautions should still be taken with this product. This product conforms to the requirements of the Safety Standards as follows: IEC :2010, EN :2010 Safety Requirements for Electrical Equipment for Measurement, Control and Laboratory use EN62471:2008 Photo-biological Safety of Lamps and Lamp Systems / Guidance on manufacturing requirements relating to non-laser optical radiation safety. Risk Group 3. This label is attached to the reverse panel of the light source. When operating or maintaining this product, please observe the following safety precautions at all times. Failure to do so may result in personal injury or damage to other items. 2.1 UV light is emitted from this product. Avoid eye and skin exposure. Never look directly into the light output beam from the LED light source. The emissions could damage the cornea and retina of the eye if the light is observed directly. 2.2 Always ensure that the LED light source is securely attached to the microscope prior to turning on the power. This will minimise the risk of injury and damage. 2.3 If for any reason the light source is to be operated when not attached to a microscope, all personnel should wear eye shielding and clothing to protect the exposed skin. DOC-005 Iss 4 4

5 2.4 Disconnecting the mains supply is achieved by unplugging the power cord from the power supply block. Only plug in the power cable, once the light source is attached to the microscope. 2.5 There are no serviceable parts within the light source. Removing any of the screws and covers will result in the safety of the light source being impaired. 2.6 Any electronic equipment connected to this product must comply with the requirements of EN/IEC To clean the exterior of the light source, use a slightly dampened cloth with a simple water/detergent solution only. Avoid the optical surfaces and lenses. Cleaning of optics should only be carried out using optical wipes and fluids. DOC-005 Iss 4 5

6 3 Getting Started System Components The CoolLED pe-300 white Illumination System is supplied with the following components: 1. LED Light Source; 2. Manual Control Pod; 3. Microscope adaptor for specific microscope model (supplied already attached to LED Light Source); 4. DC Power Supply Type GS120 A12; 5. IEC Power Cable (not shown); 6. User Guide (not shown). If any components are missing or appear damaged, please contact CoolLED immediately. DOC-005 Iss 4 6

7 4 Installation and setup 4.1 Carefully unpack the components from the shipping cartons. 4.2 Remove the protective cap from the end of the pod cable connector. 4.3 Insert the pod cable into the LED head using the red dots as a guide for orientation of the plug. 4.4 Connect the power connector from the DC power supply as shown. Ensure that the DC power supply is the one supplied with the product. Using non CoolLED power supplies may damage the light source and will invalidate the warranty. At this stage do not connect the mains power lead to the DC power supply. DOC-005 Iss 4 7

8 4.5 Attach the LED Light Source to the epi-fluorescence port on your microscope. Your pe-300 white will have been supplied with a compatible fitting for the microscope you specified at order. Attach the Light Source ensuring that it is secure and true/flush with the microscope. 4.6 Ensure that there is free airflow around the LED Light Source so that the cooling system is not impaired. A gap of 200mm on either side is sufficient. The diagram shows the Light Source in the preferred orientation. However it may be set with the cables at the top or at either side. 4.7 With the LED Light Source now attached to the microscope it is safe to connect the mains power. Connect the mains lead supplied to a convenient socket, plug in the IEC connector into the DC power supply and switch the power on at the socket. DOC-005 Iss 4 8

9 5 Configuration of LEDs as a White Light Source 5.1 Conventional white illumination systems used for fluorescence microscopy (e.g. mercury lamps), have a single element which emits light in a series of peaks across the spectrum, giving the effect of white light. LEDs are different in that a single LED element will emit light in a particular colour. To create a white illumination system, LEDs of different wavelengths have to be combined together. Using a pumped phosphor, a broader peak covering green, yellow and red emissions can also be created. In the pe-300 white, LEDs emitting in the UV and blue regions are combined with a pumped phosphor to create a white illumination system covering all the commonly used fluorescence stains. 5.2 The pe-300 white has independent circuits giving the user control of the three main peaks of emissions. On the standard configuration, these are referred to as 1UV, 2B (blue) and 3GR (green, yellow, red). DOC-005 Iss 4 9

10 5.3 There is a special variant of the pe-300 white which has been configured for use with multi-band filter sets where the first peak has been shifted from the UV region (1UV) to the violet (1V). See Appendix 1 for more information. DOC-005 Iss 4 10

11 6 Operation Manual Control 6.1 Manual Control Pod Operation on/off The pe-300 white is easily controlled from the manual control Pod. LEDs are switched on and off by pressing the on/off button. 6.2 Start-up menu. At start-up the Light Source will revert to the same settings that were set when it was last powered down. New Light Sources are supplied with the settings as shown. 6.3 To switch on LEDs press on/off once. 6.4 To switch off the LEDs, press the on/off once again. DOC-005 Iss 4 11

12 6.5 Intensity Control. The control pod enables the user to control the intensity of the LEDs that are exciting different stains. This helps to balance the emissions so that one stain does not dominate another. This feature is very useful in multi-band work (see application note in Appendix 1). 6.6 Reduce intensity of one stain by pressing the down intensity button 6.7 Individual bands can be switched off (de-selected) by pressing the select button. Light is then only generated where it is required to excite the stains in use. This has many attractive benefits with improvements in contrast, cell viability and savings in energy. Switching off UV will help to reduce damage to cells through photobleaching. DOC-005 Iss 4 12

13 6.8 Switch off selected bands by pressing the on/off button. 6.9 Switch selected bands back on by pressing on/off button again. DOC-005 Iss 4 13

14 7 Remote Operation TTL and USB 7.1 The pe-300 white can be controlled remotely via a TTL signal or through software via the USB interface. 7.2 TTL control uses the single BNC socket on the reverse of the Light Source. 7.3 The TTL signal controls the on/off function of the Light Source. A TTL high will cause the LEDs to be on, independent of the state of the on/off button. Only those bands which have been manually selected on the control Pod will be switched by the TTL signal. The intensities of the selected bands are manually set on the control pod. 7.4 The TTL input circuit has been designed to maximise the switching speed of the LEDs to give the user precise control of the excitation light reaching the sample. DOC-005 Iss 4 14

15 7.5 With fast repetitive switching, the Control pod display will not be able to respond at the same speed and so can get out of synch. If after a train of pulsing, the display on the Control pod indicates that the LEDs are on while they are actually off, simply press the on/off button to reset the display correctly. DOC-005 Iss 4 15

16 7.6 For remote control using software connection between the host computer and the pe-300 white uses the USB interface with the Light Source having a Type B connector socket located adjacent to the Control Pod socket. 7.7 Connect the Light Source to your computer using a USB cable. As with all USB remotely controlled devices, it will be necessary to set up the driver files on your system to allow the pe-300 white to be recognised. 7.8 When you first plug your CoolLED system into your PC with the USB cable, Windows will ask for a driver file unless one has already been installed. You should point Windows to the file available from CoolLED. 7.9 If you do not have the driver file you can download this from the following page on the CoolLED website: Click on the CoolLED tab near the bottom of the page and you will see the link CoolLED pe Driver. Just click on this link to download then unzip before pointing Windows to this file Once the CoolLED device has been successfully installed into Windows you should look at the Virtual COM ports assigned by going into Device Manager. Look within Ports (COM & LPT). DOC-005 Iss 4 16

17 In this example the pe-300 white has been assigned two COM ports, COM3 and COM4. You may need this information to connect to the light source from your software control package. Either COM port may be used for control. Two COM ports have been assigned to allow for diagnostics to take place in parallel with communication and also allow for dual communication should that ever be desired The majority of microscopy imaging software systems have integrated the pe-300 white into their packages. If you are developing your own software, a Software Development Kit (SDK) is available giving the full instruction sets necessary. Contact support@coolled.com and request access to this information. DOC-005 Iss 4 17

18 8 Optical setup 8.1 The pe-300 white has been designed to work on the majority of fluorescence microscopes, both new and old. As would be expected, there is some variation in the optical path and elements within every microscope. In order to accommodate these variations, the pe-300 white is supplied with a small adjustment which allows the user to optimise the performance of the illumination system when it is first fitted. This is a one-time adjustment. No further adjustment will be required during the life of the product unless changes are made to the microscope or the illumination system is fitted to a different microscope. 8.2 To make the adjustment, set up a typical sample on the microscope that gives an image over the whole field of view. Loosen the thumbscrew and slide the post back and forth until you achieve the maximum brightness with DOC-005 Iss 4 18

19 an even field of view. Tighten the thumbscrew to prevent the setting from changing. 9 Settings / Additional Information 9.1 Display Backlight and Contrast settings. The Control pod display settings can be adjusted to suit the lighting environment that the instrument is being operated in. To make adjustments, press and hold the mode button for 3 seconds. The following display will appear on the Control pod screen. Use the first column up/down buttons to adjust the backlight to the desired level. Use the second column up/down buttons to adjust the display contrast as required. DOC-005 Iss 4 19

20 To return to the main screen, either press and hold the mode button for 3 seconds again or wait 10 seconds for the screen to automatically return. 9.2 System Information To interrogate the product on its hardware and firmware revisions, press and hold the mode button for 3 seconds. Once the display settings screen appears as in 9.1, release the mode button and then give it a second short duration press. The following display will appear. To return to the main screen, either press and hold the Mode button for 3 seconds or wait for 10 seconds for the screen to automatically return. 9.3 LED hours usage The system automatically records the total time that the LEDs are actually on. To retrieve this information, repeat the process in 9.2 above except give two short duration presses of the mode button rather than a single one. The following screen will appear. DOC-005 Iss 4 20

21 To return to the main screen, either press and hold the mode button for 3 seconds or wait for 10 seconds for the screen to automatically return. 10 Troubleshooting The pe-300 white is a relatively simple system and is easy to operate. The following is a checklist should the product not operate as expected Symptom: Light Source does not power up, nothing appears on the control pod screen. Check: Is the power connector fully inserted in the pe-300 white Light Source? Is the mains lead fully inserted in the DC power supply? Is the mains supply switched on? 10.2 Symptom: Display does not respond to any button presses. Check: Is the Control pod connector fully inserted in the pe-300 white Light Source? Was the Control pod connector inserted after power up? If so, power down and power up again once Control pod connector is fully inserted to reboot the system Symptom: LEDs fail to switch on with message on screen Hot Lams. Check: Check: LEDs are over heating so check that there is sufficient clearance for airflow around the pe-300 white Light Source (see section 4.6). Check that the ambient temperature is below the maximum operating temperature (see section 14.4). Check that there are no local heat sources close to the Light Source. Listen for the fan switching on. It should not run continuously - only when deemed necessary by the system s thermal management controls. If the fan does not operate with all the LEDs on and set at maximum intensity, then there is an internal fault and the Illumination System must be returned to CoolLED for repair. (see section 16) 10.4 Symptom: Illumination appears to be weak Check: Are the intensity settings on the Control pod turned up? Has the single optical setup procedure for the pe-300 white DOC-005 Iss 4 21

22 been followed (see section 8)? Is the microscope set up correctly? Check for shutters and apertures being open and for appropriate filters and cubes. If microscope previously used a mercury or metal halide lamp, check for damage or frosting of optics in light path Symptom: Illumination on sample is not flat and even. Check: Has the single setup procedure been followed (see section 8)? 10.6 Symptom: The Illumination is not centralised over the field of view Check: Carefully slacken off the fixing retaining the pe-300 white on to the epi-port and rotate the Light Source while viewing the field of view. If the illumination offset follows the rotation, then the optics within the pe-300 white have lost their alignment, possibly through mis-handling and the product will need to be returned to CoolLED for re-aligning. If the illumination offset remains unchanged when rotating the light source then the problem is within the microscope and a person competent with microscope servicing will need to investigate the cause of the problem. DOC-005 Iss 4 22

23 11 Routine Care and Maintenance 11.1 The pe-300 white requires little or no maintenance throughout its life. There are no field serviceable parts so there is no need to remove the covers Cleaning of the external surfaces can be carried out with a mild soap and water used to lightly dampen a lint-free cloth. Ensure that no liquid is allowed to enter the product through vents and panel edges. Avoid optical surfaces Cleaning of optical surfaces maybe necessary if debris or finger prints accidently come into contact with the lens during installation. In the first instance remove any loose debris with an air duster (aerosol or rubber blower) Finger prints or other liquid type contaminants should be removed using standard lens cleaning procedures. Do not flood the lens surfaces with fluid as liquid could enter the product and cause damage. DOC-005 Iss 4 23

24 12 Fitting pe-300 white to different microscope 12.1 The pe-300 white can be easily fitted to most fluorescence microscopes, both new and old. Every microscope manufacturer has one or a number of methods of attaching the fluorescence light source. CoolLED has designed a comprehensive range of adaptors to match these microscopes There are a small number of microscopes which require additional optics or special settings internal to the pe-300 white. Light Sources for these microscopes will be supplied with a label on the back panel, next to the serial number. These light sources cannot be transferred on to other microscopes without first returning them to CoolLED for internal modifications. Contact info@coolled.com if an light source needs this modification and ensure that the complete Illumination System is returned The adaptor can be removed and replaced by simply unscrewing a pair of M4 grub screws as shown Fit the new adaptor and tighten grub screws A complete list of adaptors can be found on the CoolLED website using the link The simple optical setup procedure will need to be followed when fitting the pe-300 white to a different microscope. See section 8. DOC-005 Iss 4 24

25 13 pe-300 white with liquid light guide option 13.1 The pe-300 white is available for use with a 3mm liquid light guide rather than using a direct attachment adaptor. This enables the Illumination System to be used on microscopes which just have a liquid light guide input, the collimating optics being an integral part of the microscope Fully insert light guide as shown and tighten up grub screw to ensure end of light guide is prevented from sliding out Do not bend the liquid light guides through sharp corners. It is recommended to ensure a minimum bend radius of 75mm. Ensure that the Light Source sits upright on a flat surface and keep the clearance of 200mm on both sides to ensure adequate airflow for the cooling system. DOC-005 Iss 4 25

26 13.4 The use of a liquid light guide will be attractive for use in electrophysiology as this allows the light source to be placed outside the Faraday cage to reduce electrical noise close up to the samples. The pe- Universal Collimator is available for these applications. See section 15 for details and order codes When using this collimator, it is important to set up the optics correctly to optimise the performance of the Illumination System. Full setup instructions are given in the separate user manual for the pe-universal Collimator. DOC-005 Iss 4 26

27 14 Product specifications 14.1 Power requirements Va.c 50/60Hz 2A 14.2 Power consumption Standby mode White (all 3 bands at 100% intensity) Two bands at 100% Single band at 100% max 2W max 80W max 63W max 38W 14.3 Dimensions Light Source Control Pod Power Supply 77mm (w) x 186mm (d) x 162mm (h) -weight 1.40kg 88mm (w) x 125mm (d) x 37mm (h) - weight 0.32kg 167mm (w) x 67mm (d) x 35mm (h) - weight 0.62kg 14.4 Environmental Operating Conditions Operating 5 35 deg C DOC-005 Iss 4 27

28 15 Product options and order codes See website for full details of product options and order codes. DOC-005 Iss 4 28

29 16 Warranty and Repairs This product is supplied with a 12 month warranty which is calculated from the date of invoicing, covering defects in materials and workmanship. It does not cover or form the basis for any claims for damages or consequential losses. In addition the warranty covering the LEDs is extended from 12 months to 36 months or 25,000 hours whichever is first. Any repair that is carried out by CoolLED Ltd, or its approved agent is warranted for 100 days. The warranty does not cover excessive wear and tear, poor handling or any fault caused by operating the product outside the advice of this User Guide. In the event of the product developing a fault, make contact with support@coolled.com and provide a brief description of the problem. It is useful to have the product serial number available. If a repair is required, a support team member will provide a RMA number to log the incident. All returned units, including warranty claims should be packed up carefully and adequately (preferably in the original packaging), and sent postage and carriage paid to the service centre as specified by the support team member. DOC-005 Iss 4 29

30 17 Contact Details CoolLED Ltd Westmarch Business Centre Andover Hampshire SP10 1NS UK Phone Online +44 (0) (Worldwide) (USA + Canada) info@coolled.com DOC-005 Iss 4 30

31 18 Appendix 1 CoolLED pe-300 white with Multi-band Filter Sets New ways of working with multiband filter sets can now be achieved using the pe-300 white. These are not possible with other broad spectrum light sources. While the pe-300 white delivers a wide spectral output covering most filters sets, its intuitive manual control pod allows the user to select and adjust the intensity in three key bands of the excitation spectrum. Working with these three bands, selected and set at 100% intensity, the illumination system can be operated as a broad spectrum white light source, replacing an existing mercury or metal halide bulb. In this mode, working procedures and filter set selections remain unchanged. The three band control feature of the pe-300 white increases the practical uses of multi-band filter sets from only providing multi-colour images to also allowing single fluorophore viewing. By simply selecting or switching off regions of the excitation spectrum, single fluorophores can be viewed in isolation or in conjunction with one or two other fluorophores on the same sample. This is possible due to LED emissions being limited in bandwidth and so adding practically no energy outside the excitation region of interest. The result is reduced background with the greater signal-to-noise ratio. In addition the three band controls of the pe-300 white allow the user to vary the brightness of the individual fluorescence stains on a multi-stained sample so that a balance can be achieved between them. This can prevent brighter stains from over powering or masking weaker ones as viewed through the eyepieces. By using a pe-300 white in conjunction with a multi-band filter set, swapping between filter cubes is no longer required. The user can select which single or combination of stain emissions is being viewed solely through the pod controls. Apart from not having to remember which filter cube is in each position there are additional benefits in having a single multi-band filter set. Capturing multi-colour images with multi-band filters and conventional broadband white light sources is practical with the use of a colour camera although the ability to balance the colours is not possible. However monochrome cameras tend to be more common in microscopy labs as they are generally cheaper and provide better sensitivity and resolution than a similar pixel numbered colour camera. For this reason a multi coloured image will generally be constructed by taking a series of sequential single colour images through a series of single band filter cubes. This sequential single band filter approach does provide images with great signal to noise. However the movement between filter cubes can introduce problems in the DOC-005 Iss 4 31

32 form of time delays and vibration. Misalignment of overlaid images (pixel shift) can also occur when using different single band sets due to the individual dichroics in each cube not being set at exactly the same angle. There are complicated solutions to these issues involving the use of excitation and emission filter wheels. By contrast the pe-300 white with a straight forward multiband set can achieve both full simultaneous multicolour imaging as well as sequential imaging without the need for any moving parts. The three controllable spectral regions of the pe-300 white are: 1. UV/Violet - For UV and Violet excited fluorophores such as DAPI, Hoechst and Calcofluor White etc; 2. Blue - For Blue excited fluorophores such as GFP, FITC, Auramine etc; 3. Green/Red - For green and red excited fluorophores such as Cy3, TRITC, TxRed and mcherry as well as Cy5. Users of triple multiband filter sets should be aware that DAPI is excited at a longer Violet wavelength (400nm) than the normal UV band (365nm). This is caused by complications for the filter manufacturers in producing a single filter with three pass bands with the lowest one at 365nm. To match these triple multiband filters sets, the pe-300 white MB should be used which has the DAPI excitation band positioned in the violet region, rather than the UV. The following is a list of available triple multiband-pass sets that can be used with the pe- 300 white Violet, Blue and GYR. Chroma: ET-DAPI-FITC-TRITC ET-DAPI-FITC-Texas-Red ET-DAPI-FITC-Cy3 Semrock: DOC-005 Iss 4 32